Aberrant extracellular matrix (ECM) remodeling is a hallmark of fibrosis. Fibrosis occurs in multiple organs systems, including the heart. Alterations in the levels, composition, and organization of ECM molecules lead to changes in cell phenotype and altered tissue mechanical properties that contribute to organ function decline. Hence, understanding the underlying mechanisms that contribute to fibrosis is essential for the development of novel therapies that can limit pathologic ECM accumulation. Our data show that the ECM protein fibronectin (FN) is an important regulator of ECM remodeling. Our published data demonstrate that the process of depositing (polymerizing) FN into the ECM regulates the deposition and stability of other ECM molecules, including collagen type I. In addition, agents that we and others have used to inhibit FN polymerization, such as the peptide pUR4, block FN deposition into the ECM, trigger the loss of FN and collagen I fibrils, enhance FN and collagen I endocytosis, and inhibit inflammation. Our data show that removal of FN and collagen I from the ECM is a coordinated process that involves MT1-MMP (membrane type 1 matrix metalloproteinase) mediated extracellular degradation, followed by 1521 integrin-mediated endocytosis. Further, our preliminary data demonstrate that 1521 endocytosis is regulated by both fibronectin polymerization and MT1-MMP. These data suggest that integrin-mediated endocytosis is a major regulator of ECM remodeling, and that agents that regulate ECM endocytosis may provide a novel approach to limiting pathologic ECM accumulation. Importantly, our in vivo data show that the FN polymerization inhibitor pUR4 effectively blocks the accumulation of ECM FN and collagen I and reduces inflammation in a mouse vascular injury model. Further, our preliminary data show that pUR4 blocks fibrosis and improves cardiac function in a mouse model of myocardial infarction (MI). These data are the first to show that FN is a major contributor to adverse ECM remodeling in the cardiovascular system, and that blocking FN deposition may be an effective strategy to limit excess accumulation of ECM and inhibit inflammation during fibrosis. In this application, we will test the hypothesis that FN polymerization and MT1-MMP play key roles in regulating ECM remodeling by controlling 21 integrin endocytosis, and that regulation of ECM endocytosis is a key mechanism that limits ECM accumulation. We will also test the hypothesis that FN polymerization regulates the development of cardiac fibrosis by modulating the pro-inflammatory accumulation of FN and collagen I. These studies may lead to the development a new therapeutic approach for limiting fibrosis, for which there are few effective treatments.